This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. For bacteria, the ability to sense and adapt to environmental change and stress is crucial to persistence and survival. Two-component system (TCS) is a fundamental mechanism utilized by bacteria as well as fungi and some plants to sense external or internal signals and make appropriate responses through regulation of gene expression. TCSs play pivotal roles in pathogenesis and/or antibiotic resistance in many pathogenic microorganisms. A paradigmatic TCS consists of two proteins: a multi-domain, multi-function sensor histidine kinase (HK) that monitors environmental signals, and a response regulator (RR) that receives the signal from the sensor kinase through phosphotransfer reactions and trigger downstream responses through, e.g., regulation of expression of certain genes. Molecular mechanisms underlying TCS signal transduction remain poorly understood. The proposed research is aimed at molecular basis of TCS signal transduction at high resolution, using the YycFG of Bacillus subtilis as a model. YycFG is an essential TCS and is specific to low G+C Gram-positive bacteria such as Staphylococcus, Streptococcus and Enterococcus, which are leading causative agents of human infections and cause diseases ranging from pneumonia to meningitis, endocarditis and bacteremia. Some of these pathogens, such as Bacillus anthracis and Clostridium botulinum, are classified as potential bioterror agents. We seek to: (i) explore the domain organization of YycG at high resolution by determination of the structure of a truncated form of YycG, (ii) define the three-dimensional architecture of YycF, and (iii) investigate the molecular interactions between YycG and YycF that are critical for phosphotransfer.